BACKGROUND Intestinal ischemia reperfusion(I/R)occurs in various diseases,such as trauma and intestinal transplantation.Excessive reactive oxygen species(ROS)accumulation and subsequent apoptotic cell death in intesti...BACKGROUND Intestinal ischemia reperfusion(I/R)occurs in various diseases,such as trauma and intestinal transplantation.Excessive reactive oxygen species(ROS)accumulation and subsequent apoptotic cell death in intestinal epithelia are important causes of I/R injury.PTEN-induced putative kinase 1(PINK1)and phosphorylation of dynamin-related protein 1(DRP1)are critical regulators of ROS and apoptosis.However,the correlation of PINK1 and DRP1 and their function in intestinal I/R injury have not been investigated.Thus,examining the PINK1/DRP1 pathway may help to identify a protective strategy and improve the patient prognosis.AIM To clarify the mechanism of the PINK1/DRP1 pathway in intestinal I/R injury.METHODS Male C57BL/6 mice were used to generate an intestinal I/R model via superior mesenteric artery occlusion followed by reperfusion.Chiu’s score was used to evaluate intestinal mucosa damage.The mitochondrial fission inhibitor mdivi-1 was administered by intraperitoneal injection.Caco-2 cells were incubated in vitro in hypoxia/reoxygenation conditions.Small interfering RNAs and overexpression plasmids were transfected to regulate PINK1 expression.The protein expression levels of PINK1,DRP1,p-DRP1 and cleaved caspase 3 were measured by Western blotting.Cell viability was evaluated using a Cell Counting Kit-8 assay and cell apoptosis was analyzed by TUNEL staining.Mitochondrial fission and ROS were tested by MitoTracker and MitoSOX respectively.RESULTS Intestinal I/R and Caco-2 cell hypoxia/reoxygenation decreased the expression of PINK1 and p-DRP1 Ser637.Pretreatment with mdivi-1 inhibited mitochondrial fission,ROS generation,and apoptosis and ameliorated cell injury in intestinal I/R.Upon PINK1 knockdown or overexpression in vitro,we found that p-DRP1 Ser637 expression and DRP1 recruitment to the mitochondria were associated with PINK1.Furthermore,we verified the physical combination of PINK1 and p-DRP1 Ser637.CONCLUSION PINK1 is correlated with mitochondrial fission and apoptosis by regulating DRP1 phosphoryl展开更多
[目的]研究饲料钙对母鼠饮水型氟染毒后子代大鼠肾细胞线粒体损伤的影响。[方法]选用健康初断乳SD雌性大鼠100只,随机分为对照组、染氟组(100 mg/L Na F)、低钙组(0.063%CaCO_3)、低钙染氟组(100 mg/L Na F+0.063%CaCO_3)和高钙染氟组(1...[目的]研究饲料钙对母鼠饮水型氟染毒后子代大鼠肾细胞线粒体损伤的影响。[方法]选用健康初断乳SD雌性大鼠100只,随机分为对照组、染氟组(100 mg/L Na F)、低钙组(0.063%CaCO_3)、低钙染氟组(100 mg/L Na F+0.063%CaCO_3)和高钙染氟组(100 mg/L Na F+7%CaCO_3);饲养3个月后,雌雄鼠合笼交配。取日龄14 d及28 d仔鼠雌雄各10只,以其肾脏细胞线粒体标志酶琥珀酸脱氢酶(SDHase)活性及脂质过氧化指标丙二醛(MDA)水平,肾脏细胞凋亡状况,线粒体分裂/融合蛋白Fis1、Drp1和Mfn2表达水平为观察指标。[结果]与染氟组相比,高钙染氟组SDHase活性升高(P<0.05),低钙染氟组SDHase活性降低(P<0.05)。与对照组相比,雌鼠各组肾脏线粒体MDA含量均升高(P<0.05)。与对照组相比,染氟各组仔鼠凋亡细胞增多;与染氟组相比,低钙染氟组凋亡细胞增多,而高钙染氟组凋亡细胞减少。与对照组相比,低钙染氟组14 d雄鼠的分裂蛋白Fis1表达升高(P<0.05);低钙染氟组和高钙染氟组28 d雄鼠的分裂蛋白Drp1升高(P<0.05)。[结论]氟中毒能够造成大鼠肾脏细胞线粒体内分裂/融合蛋白Fis1、Drp1和Mfn2表达异常,引起肾脏细胞线粒体损伤。高钙饲料摄入能降低线粒体内脂质过氧化反应,减轻高氟对子代肾脏细胞的毒性作用,而低钙饲料摄入会加剧高氟的毒性作用。展开更多
目的探讨糖原合成酶激酶3β(glycogen synthase kinase 3β,GSK3β)/线粒体分裂蛋白1(fission protein 1,Fis1)信号通路在甲基乙二醛(methylglyoxal,MG)诱导成骨细胞凋亡中的作用及机制。方法采用LiCl作为GSK3β抑制剂,将细胞随机分为4...目的探讨糖原合成酶激酶3β(glycogen synthase kinase 3β,GSK3β)/线粒体分裂蛋白1(fission protein 1,Fis1)信号通路在甲基乙二醛(methylglyoxal,MG)诱导成骨细胞凋亡中的作用及机制。方法采用LiCl作为GSK3β抑制剂,将细胞随机分为4组,即对照组、MG组、LiCl组和LiCl+MG组。采用MTT法检测细胞增殖活性,Tunel染色法分析细胞凋亡情况,Western blot法检测GSK3β、Fis1蛋白表达水平,MitoTracker Deep Red染色法分析线粒体形态。结果MTT法检测结果表明,MG抑制了成骨细胞增殖活性。Tunel染色法检测结果显示,MG诱导成骨细胞凋亡。Western blot法检测结果表明,MG处理后GSK3β蛋白磷酸化水平降低,Fis1蛋白表达水平增加。MitoTracker Deep Red染色法分析结果显示,MG处理后线粒体呈碎片化。在加入GSK3β抑制剂LiCl干预后,与MG组比较,其显著恢复了MG抑制的细胞增殖活性、减少细胞凋亡,同时GSK3β蛋白磷酸化水平升高,Fis1蛋白表达水平降低,并且恢复了线粒体形态。结论MG可能通过调控GSK3β/Fis1信号通路促进线粒体分裂增加,诱导成骨细胞凋亡。展开更多
Peroxisomes and mitochondria are metabolically diverse organelles that act in concert in a number of pathways in eukaryotes, including photorespiration and lipid mobilization in plants. The division machineries of the...Peroxisomes and mitochondria are metabolically diverse organelles that act in concert in a number of pathways in eukaryotes, including photorespiration and lipid mobilization in plants. The division machineries of these two types of organelles also share several components such as dynamin-related proteins （DRPs） and their organelle anchor, the FISSION1 （FIS1） protein. In Arabidopsis, members of the DRP3 and FIS1 small protein families, namely DRP3A, DRP3B, FISIA, and FISIB, are each dual-targeted to peroxisomes and mitochondria and are required for the division of both organelles; DRP3A and DRP3B are partially redundant in function. To further determine the contribution of FISIA and FISIB to the division of peroxisomes and mitochondria, we analyzed plants overexpressing FISIA or FISIB and mutants in which the functions of both proteins were disrupted. Domains in FISIA and FISIB required for peroxisomal targeting were also dissected. Our results demonstrate that FISIA and FISIB play rate-limiting and partially overlapping roles in promoting the fission of peroxisomes and mitochondria. Furthermore, although the C-terminus of FIS1 is both necessary and sufficient for targeting to peroxisomes, the role of the short C-terminal segment adjacent to the transmembrane domain may differ among diverse species in peroxisomal targeting.展开更多
基金the National Natural Science Foundation of China,No.81679154,No.81871547.
文摘BACKGROUND Intestinal ischemia reperfusion(I/R)occurs in various diseases,such as trauma and intestinal transplantation.Excessive reactive oxygen species(ROS)accumulation and subsequent apoptotic cell death in intestinal epithelia are important causes of I/R injury.PTEN-induced putative kinase 1(PINK1)and phosphorylation of dynamin-related protein 1(DRP1)are critical regulators of ROS and apoptosis.However,the correlation of PINK1 and DRP1 and their function in intestinal I/R injury have not been investigated.Thus,examining the PINK1/DRP1 pathway may help to identify a protective strategy and improve the patient prognosis.AIM To clarify the mechanism of the PINK1/DRP1 pathway in intestinal I/R injury.METHODS Male C57BL/6 mice were used to generate an intestinal I/R model via superior mesenteric artery occlusion followed by reperfusion.Chiu’s score was used to evaluate intestinal mucosa damage.The mitochondrial fission inhibitor mdivi-1 was administered by intraperitoneal injection.Caco-2 cells were incubated in vitro in hypoxia/reoxygenation conditions.Small interfering RNAs and overexpression plasmids were transfected to regulate PINK1 expression.The protein expression levels of PINK1,DRP1,p-DRP1 and cleaved caspase 3 were measured by Western blotting.Cell viability was evaluated using a Cell Counting Kit-8 assay and cell apoptosis was analyzed by TUNEL staining.Mitochondrial fission and ROS were tested by MitoTracker and MitoSOX respectively.RESULTS Intestinal I/R and Caco-2 cell hypoxia/reoxygenation decreased the expression of PINK1 and p-DRP1 Ser637.Pretreatment with mdivi-1 inhibited mitochondrial fission,ROS generation,and apoptosis and ameliorated cell injury in intestinal I/R.Upon PINK1 knockdown or overexpression in vitro,we found that p-DRP1 Ser637 expression and DRP1 recruitment to the mitochondria were associated with PINK1.Furthermore,we verified the physical combination of PINK1 and p-DRP1 Ser637.CONCLUSION PINK1 is correlated with mitochondrial fission and apoptosis by regulating DRP1 phosphoryl
文摘[目的]研究饲料钙对母鼠饮水型氟染毒后子代大鼠肾细胞线粒体损伤的影响。[方法]选用健康初断乳SD雌性大鼠100只,随机分为对照组、染氟组(100 mg/L Na F)、低钙组(0.063%CaCO_3)、低钙染氟组(100 mg/L Na F+0.063%CaCO_3)和高钙染氟组(100 mg/L Na F+7%CaCO_3);饲养3个月后,雌雄鼠合笼交配。取日龄14 d及28 d仔鼠雌雄各10只,以其肾脏细胞线粒体标志酶琥珀酸脱氢酶(SDHase)活性及脂质过氧化指标丙二醛(MDA)水平,肾脏细胞凋亡状况,线粒体分裂/融合蛋白Fis1、Drp1和Mfn2表达水平为观察指标。[结果]与染氟组相比,高钙染氟组SDHase活性升高(P<0.05),低钙染氟组SDHase活性降低(P<0.05)。与对照组相比,雌鼠各组肾脏线粒体MDA含量均升高(P<0.05)。与对照组相比,染氟各组仔鼠凋亡细胞增多;与染氟组相比,低钙染氟组凋亡细胞增多,而高钙染氟组凋亡细胞减少。与对照组相比,低钙染氟组14 d雄鼠的分裂蛋白Fis1表达升高(P<0.05);低钙染氟组和高钙染氟组28 d雄鼠的分裂蛋白Drp1升高(P<0.05)。[结论]氟中毒能够造成大鼠肾脏细胞线粒体内分裂/融合蛋白Fis1、Drp1和Mfn2表达异常,引起肾脏细胞线粒体损伤。高钙饲料摄入能降低线粒体内脂质过氧化反应,减轻高氟对子代肾脏细胞的毒性作用,而低钙饲料摄入会加剧高氟的毒性作用。
文摘目的探讨糖原合成酶激酶3β(glycogen synthase kinase 3β,GSK3β)/线粒体分裂蛋白1(fission protein 1,Fis1)信号通路在甲基乙二醛(methylglyoxal,MG)诱导成骨细胞凋亡中的作用及机制。方法采用LiCl作为GSK3β抑制剂,将细胞随机分为4组,即对照组、MG组、LiCl组和LiCl+MG组。采用MTT法检测细胞增殖活性,Tunel染色法分析细胞凋亡情况,Western blot法检测GSK3β、Fis1蛋白表达水平,MitoTracker Deep Red染色法分析线粒体形态。结果MTT法检测结果表明,MG抑制了成骨细胞增殖活性。Tunel染色法检测结果显示,MG诱导成骨细胞凋亡。Western blot法检测结果表明,MG处理后GSK3β蛋白磷酸化水平降低,Fis1蛋白表达水平增加。MitoTracker Deep Red染色法分析结果显示,MG处理后线粒体呈碎片化。在加入GSK3β抑制剂LiCl干预后,与MG组比较,其显著恢复了MG抑制的细胞增殖活性、减少细胞凋亡,同时GSK3β蛋白磷酸化水平升高,Fis1蛋白表达水平降低,并且恢复了线粒体形态。结论MG可能通过调控GSK3β/Fis1信号通路促进线粒体分裂增加,诱导成骨细胞凋亡。
基金This work was supported by grants from the US Department of Energy and the National Science Foundation （MCB 0618335） to J.H.We would like to thank Dr Sheng Quan for cloning of the 355：：FISIA construct, Marlene Cameron for graphic assistance and Karen Bird for manuscript editing. No conflict of interest declared.
文摘Peroxisomes and mitochondria are metabolically diverse organelles that act in concert in a number of pathways in eukaryotes, including photorespiration and lipid mobilization in plants. The division machineries of these two types of organelles also share several components such as dynamin-related proteins （DRPs） and their organelle anchor, the FISSION1 （FIS1） protein. In Arabidopsis, members of the DRP3 and FIS1 small protein families, namely DRP3A, DRP3B, FISIA, and FISIB, are each dual-targeted to peroxisomes and mitochondria and are required for the division of both organelles; DRP3A and DRP3B are partially redundant in function. To further determine the contribution of FISIA and FISIB to the division of peroxisomes and mitochondria, we analyzed plants overexpressing FISIA or FISIB and mutants in which the functions of both proteins were disrupted. Domains in FISIA and FISIB required for peroxisomal targeting were also dissected. Our results demonstrate that FISIA and FISIB play rate-limiting and partially overlapping roles in promoting the fission of peroxisomes and mitochondria. Furthermore, although the C-terminus of FIS1 is both necessary and sufficient for targeting to peroxisomes, the role of the short C-terminal segment adjacent to the transmembrane domain may differ among diverse species in peroxisomal targeting.
